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| Status |
Public on Aug 23, 2020 |
| Title |
A Uromodulin mutation resulting in kidney fibrosis due to activation of the innate immune system through mononuclear phagocyte ATF4/integrated stress response signaling |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Background: Uromodulin (Tamm-Horsfall) protein is secreted by the thick ascending limb into urine and crosses the basolateral membrane into the kidney interstitium and blood. Uromodulin may engage the kidney’s innate immune system with pro and anti-inflammatory effects described. Mutations in the UMOD gene are a cause of autosomal dominant tubulointerstitial kidney disease (ADTKD) due to protein misfolding and endoplasmic reticulum stress. We identified a family with a UMOD mutation (C106F) that leads to glomerular and interstitial inflammation atypical for ADTKD.
Methods: To determine if the observed phenotype is due to mutant uromodulin induced changes in the innate or adaptive immune system, we developed a transgenic (tg) mouse with a homologous cysteine to phenylalanine mutation (C105F) in the UMOD gene using CRISPR-Cas9 gene editing.
Results: LLC-PK1 cells expressing wt and mutant protein had increased basolateral secretion of protein compared with cells expressing wt or mutant protein alone. EM examination of mutant medium showed protein aggregates in contrast to filaments in wt medium. Immunoprecipitation of plasma uromodulin from mutant mice demonstrated aggregate formation not seen in wt mice. Tg/+ mice displayed increased F4/80+ mononuclear phagocyte (MP) and podocyte inflammasome activity at baseline. Tg/+ mice developed glomerular and interstitial matrix deposition, myofibroblast proliferation and increased Creatinine with aging. Following ischemia-reperfusion injury, tg/+ mice had no increase in inflammasome activation over baseline in contrast to wt littermates and displayed improved tubular repair and renal function at 7 days. Differential gene expression analysis by RNA sequencing of kidney MP revealed activation of the ATF4 mediated stress response pathway and genes associated with the development of tissue fibrosis.
Conclusions: The C106F mutation results in glomerular and interstitial inflammatory kidney disease due to interaction of aggregated misfolded protein with mononuclear phagocytic cells resulting in ER stress response. The resulting alteration in the immune system leads to improved kidney repair following acute injury but fibrosis and chronic renal failure with aging.
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| Overall design |
To determine if changes in macrophage gene expression is a mechanism linking phagocytosis of aggregated uromodulin to the pathogenesis of disease in transgenic mice, we performed RNA-sequencing of mRNA isolated from F4/80+ cells purified from 4 mutant mice and 6 wt littermate controls at 12-16 weeks using anti-F4/80 conjugated magnetic microbeads. Total N=10.
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| Contributor(s) |
Plotkin M, Obrien C, Goellner J, Williams J, Carter W, Stone A |
| Citation(s) |
32926855 |
| |
| Submission date |
Oct 30, 2019 |
| Last update date |
Sep 17, 2020 |
| Contact name |
Annjanette Stone |
| E-mail(s) |
[email protected], [email protected]
|
| Phone |
501-257-5178
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| Organization name |
CAVHS
|
| Department |
Research
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| Lab |
Pharmacogenomics Analysis Laboratory
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| Street address |
4300 West 7th Street
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| City |
Little Rock |
| State/province |
AR |
| ZIP/Postal code |
72205 |
| Country |
USA |
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| Platforms (1) |
| GPL24247 |
Illumina NovaSeq 6000 (Mus musculus) |
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| Samples (10)
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| Relations |
| BioProject |
PRJNA580376 |
| SRA |
SRP227445 |
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